S271: BONE MARROW TFR2 DELETION IMPROVES THE THERAPEUTIC EFFECT OF ACTIVIN LIGAND TRAP RAP-536 IN Β-THALASSEMIC MICE
Authors: E. Tanzi, S. M Di Modica, J. Bordini, V. Olivari, M. Pettinato, A. Pagani, A. Campanella, L. Silvestri, A. Nai
Journal: HemaSphere (2022)

Background: β-thalassemia is a disorder due to mutations in the β-globin gene, characterized by ineffective erythropoiesis (IE), anemia, and iron overload, whose treatment still requires improvement. Luspatercept, an activin receptor-ligand trap that promotes terminal erythroid differentiation downregulating the TGF-β pathway is a recently approved drug for treatment of transfusion dependent β-thalassemia (TDT) and in phase 3 trial for non TDT (NTDT). However, its precise mechanism of action remains to be elucidated and the possible connection with the erythropoiesis driving cytokine erythropoietin (EPO) is still controversial. In addition, Luspatercept is only partially effective in correcting anemia, thus the identification of strategies for enhancing its efficacy would be of clinical benefit. The second transferrin receptor (TFR2) balances red blood cells (RBC) production with iron availability, contributing to the activation of the iron hormone hepcidin in the liver and acting as an EPO receptor partner in erythroid cells. We have demonstrated that bone marrow (BM) Tfr2 deletion increases erythroblast EPO sensitivity causing erythrocytosis in wild-type mice (Nai et al, Blood 2015) and improving IE, anemia, RBCs morphology and iron-overload in NTDT mice (Artuso et al, Blood 2018). Aims: As we have previously demonstrated that the combined targeting of different pathophysiologic cues of the disease results in a synergistic benefit on thalassemic erythropoiesis and anemia (Casu, Pettinato, et al, Blood 2020), here we investigate whether BM Tfr2 genetic inactivation improves the efficacy of Luspatercept in a NTDT model (Hbbth3/+). Methods:Hbbth3/+ mice with BM Tfr2 deletion (Tfr2BMKO/Hbbth3/+) and Hbbth3/+ controls were treated with RAP-536 (Celgene, BMS group), a murine analogue of Luspatercept, 10mg/kg, twice a week for 8 weeks. Complete blood count was evaluated every two weeks. At sacrifice, blood and organs were collected for full phenotypic analysis. Results: As expected, BM Tfr2 deficiency strongly ameliorated anemia and IE of Hbbth3/+ mice (hemoglobin, Hb=13.48±0.19g/dL vs 8.65±0.24g/dL in controls, P<0.000001). RAP-536 moderately increased RBC count, Hb and hematocrit after 2 weeks of treatment, and the beneficial effect was maintained until the end of the protocol with a similar efficacy in Hbbth3/+ (Hb=9.54±0.33g/dL) and Tfr2BMKO/Hbbth3/+ mice (Hb=14.48±0.50g/dL), without significant changes in MCV, MCH and reticulocyte count. Splenomegaly was not affected by RAP-536 in Hbbth3/+ mice, despite an amelioration of both spleen and BM IE, with reduced proportion of immature erythroblasts and increased mature RBCs. Erythroid differentiation was further improved in RAP-536-treated Tfr2BMKO/Hbbth3/+ mice both in the spleen and in the BM, leading to a significant reduction of spleen size, to values of wt animals. Hepatic and serum iron levels were reduced in mice lacking Tfr2, likely because of high erythroid consumption, while RAP-536 did not affect iron homeostasis in both genotypes. Summary/Conclusion: BM Tfr2 deletion and RAP-536 treatment had additive effect in correcting IE and anemia in β-thalassemic mice. Since RAP-536 has comparable therapeutic efficacy in Hbbth3/+ and Tfr2BMKO/Hbbth3/+ animals, it improves erythropoiesis through mechanism(s) other than Tfr2 deletion, thus independent of EPO stimulation. Overall, our results add a piece of information on the mechanism of action of Luspatercept and suggest that the concomitant TFR2 targeting might represent a promising option for boosting the therapeutic efficacy of activin receptor ligand-traps.